ABSTRACT A genetic haplotype, consisting of two amino acid substitutions, in the gene encoding complement factor H (CFH) significantly increases the risk for age-related macular degeneration (AMD), the leading cause of blindness in developed countries. The role of CFH, and in particular its AMD-linked haplotype in the cellular and molecular mechanisms underlying AMD pathogenesis remains largely unknown. This project proposes to utilize CRISPR-Cas9 genome editing to establish stable cell lines of human retinal pigment epithelium (RPE), expressing genetic variants of CFH, including the AMD-linked variant. Given that the RPE is a major source of CFH in the retina, and has been reported to be a primary site of insult in AMD, these RPE cell lines will serve as a valid in vitro model to test the central hypothesis that the AMD-linked haplotype compromises CFH?s function and, ultimately, results in an RPE phenotype. Specifically, the model will be used to test the role of CFH in several homeostatic functions and events in the RPE, including barrier function, tolerance to oxidative stress, degradation of photoreceptor outer segments (POS), and formation of sub-RPE deposits. Sensitive live- cell assays, including four-dimensional live-cell imaging and transepithelial resistance measurement, will be used to test POS degradation and barrier function, respectively. In summary, this project will address key questions in human RPE cell biology ? (1) Does the AMD-linked haplotype of CFH compromise its function in the RPE? (2) What RPE cellular phenotypes result from compromised CFH function? (3) What endogenous role does CFH plays in maintaining RPE homeostasis?